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18 pages, 2709 KiB

Open AccessArticle

Photoreactivity of Hair Melanin from Different Skin Phototypes—Contribution of Melanin Subunits to the Pigments Photoreactive Properties

by Krystian Mokrzynski, Shosuke Ito, Kazumasa Wakamatsu, Theodore G. Camenish, Tadeusz Sarna and Michal Sarna

Int. J. Mol. Sci. 2021, 22(9), 4465; https://doi.org/10.3390/ijms22094465 - 24 Apr 2021

Cited by 5 | Viewed by 3102

Abstract

Photoreactivity of melanin has become a major focus of research due to the postulated involvement of the pigment in UVA-induced melanoma. However, most of the hitherto studies were carried out using synthetic melanin models. Thus, photoreactivity of natural melanins is yet to be [...] Read more.

Photoreactivity of melanin has become a major focus of research due to the postulated involvement of the pigment in UVA-induced melanoma. However, most of the hitherto studies were carried out using synthetic melanin models. Thus, photoreactivity of natural melanins is yet to be systematically analyzed. Here, we examined the photoreactive properties of natural melanins isolated from hair samples obtained from donors of different skin phototypes (I, II, III, and V). X-band and W-band electron paramagnetic resonance (EPR) spectroscopy was used to examine the paramagnetic properties of the pigments. Alkaline hydrogen peroxide degradation and hydroiodic acid hydrolysis were used to determine the chemical composition of the melanins. EPR oximetry and spin trapping were used to examine the oxygen photoconsumption and photo-induced formation of superoxide anion, and time-resolved near infrared phosphorescence was employed to determine the singlet oxygen photogeneration by the melanins. The efficiency of superoxide and singlet oxygen photogeneration was related to the chemical composition of the studied melanins. Melanins from blond and chestnut hair (phototypes II and III) exhibited highest photoreactivity of all examined pigments. Moreover, melanins of these phototypes showed highest quantum efficiency of singlet oxygen photogeneration at 332 nm and 365 nm supporting the postulate of the pigment contribution in UVA-induced melanoma. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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15 pages, 2415 KiB

Open AccessArticle

Density Functional Theory-Based Calculation Shed New Light on the Bizarre Addition of Cysteine Thiol to Dopaquinone

by Ryo Kishida, Shosuke Ito, Manickam Sugumaran, Ryan Lacdao Arevalo, Hiroshi Nakanishi and Hideaki Kasai

Int. J. Mol. Sci. 2021, 22(3), 1373; https://doi.org/10.3390/ijms22031373 - 29 Jan 2021

Cited by 9 | Viewed by 2577

Abstract

Two types of melanin pigments, brown to black eumelanin and yellow to reddish brown pheomelanin, are biosynthesized through a branched reaction, which is associated with the key intermediate dopaquinone (DQ). In the presence of l-cysteine, DQ immediately binds to the –SH group, [...] Read more.

Two types of melanin pigments, brown to black eumelanin and yellow to reddish brown pheomelanin, are biosynthesized through a branched reaction, which is associated with the key intermediate dopaquinone (DQ). In the presence of l-cysteine, DQ immediately binds to the –SH group, resulting in the formation of cysteinyldopa necessary for the pheomelanin production. l-Cysteine prefers to bond with aromatic carbons adjacent to the carbonyl groups, namely C5 and C2. Surprisingly, this Michael addition takes place at 1,6-position of the C5 (and to some extent at C2) rather than usually expected 1,4-position. Such an anomaly on the reactivity necessitates an atomic-scale understanding of the binding mechanism. Using density functional theory-based calculations, we investigated the binding of l-cysteine thiolate (Cys–S⁻) to DQ. Interestingly, the C2–S bonded intermediate was less energetically stable than the C6–S bonded case. Furthermore, the most preferred Cys–S⁻-attacked intermediate is at the carbon-carbon bridge between the two carbonyls (C3–C4 bridge site) but not on the C5 site. This structure allows the Cys–S⁻ to migrate onto the adjacent C5 or C2 with small activation energies. Further simulation demonstrated a possible conversion pathway of the C5–S (and C2–S) intermediate into 5-S-cysteinyldopa (and 2-S-cysteinyldopa), which is the experimentally identified major (and minor) product. Based on the results, we propose that the binding of Cys–S⁻ to DQ proceeds via the following path: (i) coordination of Cys–S⁻ to C3–C4 bridge, (ii) migration of Cys–S⁻ to C5 (C2), (iii) proton rearrangement from cysteinyl –NH₃⁺ to O4 (O3), and (iv) proton rearrangement from C5 (C2) to O3 (O4). Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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16 pages, 1775 KiB

Open AccessArticle

Chemical Evaluation of Eumelanin Maturation by ToF-SIMS and Alkaline Peroxide Oxidation HPLC Analysis

by Martin Jarenmark, Peter Sjövall, Shosuke Ito, Kazumasa Wakamatsu and Johan Lindgren

Int. J. Mol. Sci. 2021, 22(1), 161; https://doi.org/10.3390/ijms22010161 - 26 Dec 2020

Cited by 8 | Viewed by 2355

Abstract

Residual melanins have been detected in multimillion-year-old animal body fossils; however, confident identification and characterization of these natural pigments remain challenging due to loss of chemical signatures during diagenesis. Here, we simulate this post-burial process through artificial maturation experiments using three synthetic and [...] Read more.

Residual melanins have been detected in multimillion-year-old animal body fossils; however, confident identification and characterization of these natural pigments remain challenging due to loss of chemical signatures during diagenesis. Here, we simulate this post-burial process through artificial maturation experiments using three synthetic and one natural eumelanin exposed to mild (100 °C/100 bar) and harsh (250 °C/200 bar) environmental conditions, followed by chemical analysis employing alkaline hydrogen peroxide oxidation (AHPO) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). Our results show that AHPO is sensitive to changes in the melanin molecular structure already during mild heat and pressure treatment (resulting, e.g., in increased C-C cross-linking), whereas harsh maturation leads to extensive loss of eumelanin-specific chemical markers. In contrast, negative-ion ToF-SIMS spectra are considerably less affected by mild maturation conditions, and eumelanin-specific features remain even after harsh treatment. Detailed analysis of ToF-SIMS spectra acquired prior to experimental treatment revealed significant differences between the investigated eumelanins. However, systematic spectral changes upon maturation reduced these dissimilarities, indicating that intense heat and pressure treatment leads to the formation of a common, partially degraded, eumelanin molecular structure. Our findings elucidate the complementary nature of AHPO and ToF-SIMS during chemical characterization of eumelanin traces in fossilized organismal remains. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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13 pages, 3229 KiB

Open AccessArticle

Nonenzymatic Spontaneous Oxidative Transformation of 5,6-Dihydroxyindole

by Manickam Sugumaran, Jason Evans, Shosuke Ito and Kazumasa Wakamatsu

Int. J. Mol. Sci. 2020, 21(19), 7321; https://doi.org/10.3390/ijms21197321 - 03 Oct 2020

Cited by 17 | Viewed by 3115

Abstract

Melanin is an important phenolic skin pigment found throughout the animal kingdom. Tyrosine and its hydroxylated product dopa provide the starting material for melanin biosynthesis in all animals. Through a set of well-established reactions, they are converted to 5,6-dihydroxyindole (DHI) and DHI-2-carboxylic acid [...] Read more.

Melanin is an important phenolic skin pigment found throughout the animal kingdom. Tyrosine and its hydroxylated product dopa provide the starting material for melanin biosynthesis in all animals. Through a set of well-established reactions, they are converted to 5,6-dihydroxyindole (DHI) and DHI-2-carboxylic acid (DHICA). Oxidative polymerization of these two indoles produces the brown to black eumelanin pigment. The steps associated with these transformations are complicated by the extreme instability of the starting materials and the transient and highly reactive nature of the intermediates. We have used mass spectral studies to explore the nonenzymatic mechanism of oxidative transformation of DHI in water. Our results indicate the facile production of not only dimeric and trimeric products but also higher oligomeric forms of DHI upon exposure to air in solution, even under nonenzymatic conditions. Such instantaneous polymerization of DHI avoids toxicity to self-matter and ensures the much-needed deposition of melanin at (a) the wound site and (b) the infection site in arthropods. The rapid deposition of DHI melanin is advantageous for arthropods given their open circulatory system; the process limits blood loss during wounding and prevents the spread of parasites by encapsulating them in melanin, limiting the damage. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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11 pages, 3781 KiB

Open AccessArticle

Oxidative Oligomerization of DBL Catechol, a potential Cytotoxic Compound for Melanocytes, Reveals the Occurrence of Novel Ionic Diels-Alder Type Additions

by Manickam Sugumaran, Kubra Umit, Jason Evans, Rachel Muriph, Shosuke Ito and Kazumasa Wakamatsu

Int. J. Mol. Sci. 2020, 21(18), 6774; https://doi.org/10.3390/ijms21186774 - 15 Sep 2020

Cited by 3 | Viewed by 3077

Abstract

The exposure of human skin to 4-(4-hydroxyphenyl)-2-butanone (raspberry ketone, RK) is known to cause chemical/occupational leukoderma. RK is a carbonyl derivative of 4-(4-hydroxyphenyl)-2-butanol (rhododendrol), a skin whitening agent that was found to cause leukoderma in skin of many consumers. These two phenolic compounds [...] Read more.

The exposure of human skin to 4-(4-hydroxyphenyl)-2-butanone (raspberry ketone, RK) is known to cause chemical/occupational leukoderma. RK is a carbonyl derivative of 4-(4-hydroxyphenyl)-2-butanol (rhododendrol), a skin whitening agent that was found to cause leukoderma in skin of many consumers. These two phenolic compounds are oxidized by tyrosinase and the resultant products seem to cause cytotoxicity to melanocytes by producing reactive oxygen species and depleting cellular thiols through o-quinone oxidation products. Therefore, it is important to understand the biochemical mechanism of the oxidative transformation of these compounds. Earlier studies indicate that RK is initially oxidized to RK quinone by tyrosinase and subsequently converted to a side chain desaturated catechol called 3,4-dihydroxybenzalacetone (DBL catechol). In the present study, we report the oxidation chemistry of DBL catechol. Using UV–visible spectroscopic studies and liquid chromatography mass spectrometry, we have examined the reaction of DBL catechol with tyrosinase and sodium periodate. Our results indicate that DBL quinone formed in the reaction is extremely reactive and undergoes facile dimerization and trimerization reactions to produce multiple isomeric products by novel ionic Diels-Alder type condensation reactions. The production of a wide variety of complex quinonoid products from such reactions would be potentially more toxic to cells by causing not only oxidative stress, but also melanotoxicity through exhibiting reactions with cellular macromolecules and thiols. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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12 pages, 2238 KiB

Open AccessArticle

UP256 Inhibits Hyperpigmentation by Tyrosinase Expression/Dendrite Formation via Rho-Dependent Signaling and by Primary Cilium Formation in Melanocytes

by Min Cheol Kang, Jae-Wook Lee, Taek Hwan Lee, Lalita Subedi, Hussain M. Wahedi, Seon-Gil Do, Eunju Shin, Eun-Yi Moon and Sun Yeou Kim

Int. J. Mol. Sci. 2020, 21(15), 5341; https://doi.org/10.3390/ijms21155341 - 28 Jul 2020

Cited by 15 | Viewed by 3921

Abstract

Skin hyperpigmentation is generally characterized by increased synthesis and deposition of melanin in the skin. UP256, containing bakuchiol, is a well-known medication for acne vulgaris. Acne sometimes leaves dark spots on the skin, and we hypothesized that UP256 may be effective against hyperpigmentation-associated [...] Read more.

Skin hyperpigmentation is generally characterized by increased synthesis and deposition of melanin in the skin. UP256, containing bakuchiol, is a well-known medication for acne vulgaris. Acne sometimes leaves dark spots on the skin, and we hypothesized that UP256 may be effective against hyperpigmentation-associated diseases. UP256 was treated for anti-melanogenesis and melanocyte dendrite formation in cultured normal human epidermal melanocytes as well as in the reconstituted skin and zebrafish models. Western blot analysis and glutathione S-transferase (GST)-pull down assays were used to evaluate the expression and interaction of enzymes related in melanin synthesis and transportation. The cellular tyrosinase activity and melanin content assay revealed that UP256 decreased melanin synthesis by regulating the expression of proteins related on melanogenesis including tyrosinase, TRP-1 and -2, and SOX9. UP256 also decreased dendrite formation in melanocytes via regulating the Rac/Cdc42/α-PAK signaling proteins, without cytotoxic effects. UP256 also inhibited ciliogenesis-dependent melanogenesis in normal human epidermal melanocytes. Furthermore, UP256 suppressed melanin contents in the zebrafish and the 3D human skin tissue model. All things taken together, UP256 inhibits melanin synthesis, dendrite formation, and primary cilium formation leading to the inhibition of melanogenesis. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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12 pages, 3312 KiB

Open AccessArticle

Improved HPLC Conditions to Determine Eumelanin and Pheomelanin Contents in Biological Samples Using an Ion Pair Reagent

by Shosuke Ito, Sandra Del Bino, Tomohisa Hirobe and Kazumasa Wakamatsu

Int. J. Mol. Sci. 2020, 21(14), 5134; https://doi.org/10.3390/ijms21145134 - 20 Jul 2020

Cited by 22 | Viewed by 3328

Abstract

Alkaline hydrogen peroxide oxidation (AHPO) of eumelanin and pheomelanin, two major classes of melanin pigments, affords pyrrole-2,3,5-tricarboxylic acid (PTCA), pyrrole-2,3-dicarboxylic acid (PDCA) and pyrrole-2,3,4,5-tetracarboxylic acid (PTeCA) from eumelanin and thiazole-2,4,5-tricarboxylic acid (TTCA) and thiazole-4,5-dicarboxylic acid (TDCA) from pheomelanin. Quantification of these five markers [...] Read more.

Alkaline hydrogen peroxide oxidation (AHPO) of eumelanin and pheomelanin, two major classes of melanin pigments, affords pyrrole-2,3,5-tricarboxylic acid (PTCA), pyrrole-2,3-dicarboxylic acid (PDCA) and pyrrole-2,3,4,5-tetracarboxylic acid (PTeCA) from eumelanin and thiazole-2,4,5-tricarboxylic acid (TTCA) and thiazole-4,5-dicarboxylic acid (TDCA) from pheomelanin. Quantification of these five markers by HPLC provides useful information on the quantity and structural diversity of melanins in various biological samples. HPLC analysis of these markers using the original method of 0.1 M potassium phosphate buffer (pH 2.1):methanol = 99:1 (85:15 for PTeCA) on a reversed-phase column had some problems, including the short lifetime of the column and, except for the major eumelanin marker PTCA, other markers were occasionally overlapped by interfering peaks in samples containing only trace levels of these markers. These problems can be overcome by the addition of an ion pair reagent for anions, such as tetra-n-butylammonium bromide (1 mM), to retard the elution of di-, tri- and tetra-carboxylic acids. The methanol concentration was increased to 17% (30% for PTeCA) and the linearity, reproducibility, and recovery of the markers with this improved method is good to excellent. This improved HPLC method was compared to the original method using synthetic melanins, mouse hair, human hair, and human epidermal samples. In addition to PTCA, TTCA, a major marker for pheomelanin, showed excellent correlations between both HPLC methods. The other markers showed an attenuation of the interfering peaks with the improved method. We recommend this improved HPLC method for the quantitative analysis of melanin markers following AHPO because of its simplicity, accuracy, and reproducibility. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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13 pages, 3518 KiB

Open AccessArticle

Tyrosinase-Targeting Gallacetophenone Inhibits Melanogenesis in Melanocytes and Human Skin- Equivalents

by Ji Young Lee, Jooyun Lee, Daejin Min, Juewon Kim, Hyoung-June Kim and Kyoung Tai No

Int. J. Mol. Sci. 2020, 21(9), 3144; https://doi.org/10.3390/ijms21093144 - 29 Apr 2020

Cited by 14 | Viewed by 3865

Abstract

Demands for safe depigmentation compounds are constantly increasing in the pharmaceutical and cosmetic industry, since the numerous relevant compounds reported to date have shown undesirable side effects or low anti-melanogenic effects. In this study, we reported three novel inhibitors of tyrosinase, which is [...] Read more.

Demands for safe depigmentation compounds are constantly increasing in the pharmaceutical and cosmetic industry, since the numerous relevant compounds reported to date have shown undesirable side effects or low anti-melanogenic effects. In this study, we reported three novel inhibitors of tyrosinase, which is the key enzyme in melanogenesis, identified using docking-based high throughput virtual screening of an in-house natural compound library followed by mushroom tyrosinase inhibition assay. Of the three compounds, gallacetophenone showed high anti-melanogenic effect in both human epidermal melanocytes and a 3D human skin model, MelanoDerm. The inhibitory effect of gallacetophenone on tyrosinase was elucidated by computational molecular modeling at the atomic level. Binding of gallacetophenone to the active site of tyrosinase was found to be stabilized by hydrophobic interactions with His367, Ile368, and Val377; hydrogen bonding with Ser380 and a water molecule bridging the copper ions. Thus, our results strongly suggested gallacetophenone as an anti-melanogenic ingredient that inhibits tyrosinase. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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17 pages, 3947 KiB

Open AccessArticle

Argania Spinosa Fruit Shell Extract-Induced Melanogenesis via cAMP Signaling Pathway Activation

by Rachida Makbal, Myra O. Villareal, Chemseddoha Gadhi, Abdellatif Hafidi and Hiroko Isoda

Int. J. Mol. Sci. 2020, 21(7), 2539; https://doi.org/10.3390/ijms21072539 - 06 Apr 2020

Cited by 18 | Viewed by 3719

Abstract

We have previously reported that argan oil and argan press-cake from the kernels of Argania spinosa have an anti-melanogenesis effect. Here, the effect of argan fruit shell ethanol extract (AFSEE) on melanogenesis in B16F10 cells was determined, and the mechanism underlying its effect [...] Read more.

We have previously reported that argan oil and argan press-cake from the kernels of Argania spinosa have an anti-melanogenesis effect. Here, the effect of argan fruit shell ethanol extract (AFSEE) on melanogenesis in B16F10 cells was determined, and the mechanism underlying its effect was elucidated. The proliferation of AFSEE-treated B16F10 cells was evaluated using the 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) assay, while the melanin content was quantified using a spectrophotometric method. The expression of melanogenesis-related proteins was determined by Western blot and real-time PCR, while global gene expression was determined using a DNA microarray. In vitro analysis results showed that the melanin content of B16F10 cells was significantly increased by AFSEE, without cytotoxicity, by increasing the melanogenic enzyme tyrosinase (TRY), tyrosinase related-protein 1 (TRP1), and dopachrome tautomerase (DCT) protein and mRNA expression, as well as upregulating microphthalmia-associated transcription factor (MITF) expression through mitogen-activated protein kinases (MAPKs) extracellular signal-regulated kinase (ERK) and p38, and the cyclic adenosine monophosphate (cAMP) signaling pathway, as indicated by the microarray analysis results. AFSEE’s melanogenesis promotion effect is primarily attributed to its polyphenolic components. In conclusion, AFSEE promotes melanogenesis in B16F10 cells by upregulating the expression of the melanogenic enzymes through the cAMP–MITF signaling pathway.AFSEE may be used as a cosmetics product component to promote melanogenesis, or as a therapeutic against hypopigmentation disorders. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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13 pages, 5050 KiB

Open AccessArticle

Molecular and Potential Regulatory Mechanisms of Melanin Synthesis in Harmonia axyridis

by Da Xiao, Xu Chen, Renbin Tian, Mengmeng Wu, Fan Zhang, Liansheng Zang, James D. Harwood and Su Wang

Int. J. Mol. Sci. 2020, 21(6), 2088; https://doi.org/10.3390/ijms21062088 - 18 Mar 2020

Cited by 7 | Viewed by 3399

Abstract

Melanization is a common phenomenon in insects, and melanin synthesis is a conserved physiological process that occurs in epidermal cells. Moreover, a comprehensive understanding of the mechanisms of melanin synthesis influencing insect pigmentation are well-suited for investigating phenotype variation. The Asian multi-colored (Harlequin) [...] Read more.

Melanization is a common phenomenon in insects, and melanin synthesis is a conserved physiological process that occurs in epidermal cells. Moreover, a comprehensive understanding of the mechanisms of melanin synthesis influencing insect pigmentation are well-suited for investigating phenotype variation. The Asian multi-colored (Harlequin) ladybird beetle, Harmonia axyridis, exhibits intraspecific polymorphism based on relative levels of melanization. However, the specific characteristics of melanin synthesis in H. axyridis remains elusive. In this study, we performed gene-silencing analysis of the pivotal inverting enzyme, tyrosine hydroxylase (TH), and DOPA decarboxylase (DDC) in the tyrosine metabolism pathway to investigate the molecular and regulatory mechanism of melanin synthesis in H. axyridis. Using RNAi of TH and DDC genes in fourth instar larvae, we demonstrated that dopamine melanin was the primary contributor to the overall body melanization of H. axyridis. Furthermore, our study provides the first conclusive evidence that dopamine serves as a melanin precursor for synthesis in the early pupal stage. According to transcription factor Pannier, which is essential for the formation of melanic color on the elytra in H. axyridis, we further demonstrated that suppression of HaPnr can significantly decrease expression levels of HaTH and HaDDC. These results in their entirety lead to the conclusion that transcription factor Pannier can regulate dopamine melanin synthesis in the dorsal elytral epidermis of H. axyridis. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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13 pages, 3960 KiB

Open AccessArticle

Glucose Exerts an Anti-Melanogenic Effect by Indirect Inactivation of Tyrosinase in Melanocytes and a Human Skin Equivalent

by Sung Hoon Lee, Il-Hong Bae, Eun-Soo Lee, Hyoung-June Kim, Jongsung Lee and Chang Seok Lee

Int. J. Mol. Sci. 2020, 21(5), 1736; https://doi.org/10.3390/ijms21051736 - 03 Mar 2020

Cited by 15 | Viewed by 5027

Abstract

Sugars are ubiquitous in organisms and well-known cosmetic ingredients for moisturizing skin with minimal side-effects. Glucose, a simple sugar used as an energy source by living cells, is often used in skin care products. Several reports have demonstrated that sugar and sugar-related compounds [...] Read more.

Sugars are ubiquitous in organisms and well-known cosmetic ingredients for moisturizing skin with minimal side-effects. Glucose, a simple sugar used as an energy source by living cells, is often used in skin care products. Several reports have demonstrated that sugar and sugar-related compounds have anti-melanogenic effects on melanocytes. However, the underlying molecular mechanism by which glucose inhibits melanin synthesis is unknown, even though glucose is used as a whitening as well as moisturizing ingredient in cosmetics. Herein, we found that glucose significantly reduced the melanin content of α-melanocyte-stimulating hormone (MSH)-stimulated B16 cells and darkly pigmented normal human melanocytes with no signs of cytotoxicity. Furthermore, topical treatment of glucose clearly demonstrated its whitening efficacy through photography, Fontana-Masson (F&M) staining, and multi-photon microscopy in a pigmented 3D human skin model, MelanoDerm. However, glucose did not alter the gene expression or protein levels of major melanogenic proteins in melanocytes. While glucose potently decreased intracellular tyrosinase activity in melanocytes, it did not reduce mushroom tyrosinase activity in a cell-free experimental system. However, glucose was metabolized into lactic acid, which can powerfully suppress tyrosinase activity. Thus, we concluded that glucose indirectly inhibits tyrosinase activity through conversion into lactic acid, explaining its anti-melanogenic effects in melanocytes. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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18 pages, 4090 KiB

Open AccessArticle

The Surprising Effect of Phenformin on Cutaneous Darkening and Characterization of Its Underlying Mechanism by a Forward Chemical Genetics Approach

by Kei Takano, Akira Hachiya, Daiki Murase, Akiko Kawasaki, Hirokazu Uda, Shinya Kasamatsu, Yoshiya Sugai, Yoshito Takahashi, Tadashi Hase, Atsushi Ohuchi and Tamio Suzuki

Int. J. Mol. Sci. 2020, 21(4), 1451; https://doi.org/10.3390/ijms21041451 - 20 Feb 2020

Cited by 3 | Viewed by 3318

Abstract

Melanin in the epidermis is known to ultimately regulate human skin pigmentation. Recently, we exploited a phenotypic-based screening system composed of ex vivo human skin cultures to search for effective materials to regulate skin pigmentation. Since a previous study reported the potent inhibitory [...] Read more.

Melanin in the epidermis is known to ultimately regulate human skin pigmentation. Recently, we exploited a phenotypic-based screening system composed of ex vivo human skin cultures to search for effective materials to regulate skin pigmentation. Since a previous study reported the potent inhibitory effect of metformin on melanogenesis, we evaluated several biguanide compounds. The unexpected effect of phenformin, once used as an oral anti-diabetic drug, on cutaneous darkening motivated us to investigate its underlying mechanism utilizing a chemical genetics approach, and especially to identify alternatives to phenformin because of its risk of severe lactic acidosis. Chemical pull-down assays with phenformin-immobilized beads were performed on lysates of human epidermal keratinocytes, and subsequent mass spectrometry identified 7-dehydrocholesterol reductase (DHCR7). Consistent with this, AY9944, an inhibitor of DHCR7, was found to decrease autophagic melanosome degradation in keratinocytes and to intensely darken skin in ex vivo cultures, suggesting the involvement of cholesterol biosynthesis in the metabolism of melanosomes. Thus, our results validated the combined utilization of the phenotypic screening system and chemical genetics as a new approach to develop promising materials for brightening/lightening and/or tanning technologies. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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13 pages, 3971 KiB

Open AccessArticle

Rhododenol Activates Melanocytes and Induces Morphological Alteration at Sub-Cytotoxic Levels

by Minjeong Kim, Chang-Seok Lee and Kyung-Min Lim

Int. J. Mol. Sci. 2019, 20(22), 5665; https://doi.org/10.3390/ijms20225665 - 12 Nov 2019

Cited by 9 | Viewed by 3278

Abstract

Rhododenol (RD), a whitening cosmetic ingredient, was withdrawn from the market due to RD-induced leukoderma (RIL). While many attempts have been made to clarify the mechanism underlying RIL, RIL has not been fully understood yet. Indeed, affected subjects showed uneven skin pigmentation, but [...] Read more.

Rhododenol (RD), a whitening cosmetic ingredient, was withdrawn from the market due to RD-induced leukoderma (RIL). While many attempts have been made to clarify the mechanism underlying RIL, RIL has not been fully understood yet. Indeed, affected subjects showed uneven skin pigmentation, but the features are different from vitiligo, a skin hypopigmentary disorder, alluding to events more complex than simple melanocyte cytotoxicity. Here, we discovered that rhododenol treatment reduced the number of melanocytes in a pigmented 3D human skin model, Melanoderm™, confirming the melanocyte toxicity of RD. Of note, melanocytes that survived in the RD treated tissues exhibited altered morphology, such as extended dendrites and increased cell sizes. Consistently with this, sub-cytotoxic level of RD increased cell size and elongated dendrites in B16 melanoma cells. Morphological changes of B16 cells were further confirmed in the immunocytochemistry of treated cells for actin and tubulin. Even more provoking, RD up-regulated the expression of tyrosinase and TRP1 in the survived B16 cells. Evaluation of mRNA expression of cytoskeletal proteins suggests that RD altered the cytoskeletal dynamic favoring cell size expansion and melanosome maturation. Collectively, these results suggest that RD not only induces cytotoxicity in melanocytes but also can lead to a profound perturbation of melanocyte integrity even at sub-cytotoxic levels. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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13 pages, 2335 KiB

Open AccessArticle

The Organogermanium Compound THGP Suppresses Melanin Synthesis via Complex Formation with L-DOPA on Mushroom Tyrosinase and in B16 4A5 Melanoma Cells

by Junya Azumi, Tomoya Takeda, Yasuhiro Shimada, Hisashi Aso and Takashi Nakamura

Int. J. Mol. Sci. 2019, 20(19), 4785; https://doi.org/10.3390/ijms20194785 - 26 Sep 2019

Cited by 15 | Viewed by 7081

Abstract

The organogermanium compound 3-(trihydroxygermyl)propanoic acid (THGP) has various biological activities. We previously reported that THGP forms a complex with cis-diol structures. L-3,4-Dihydroxyphenylalanine (L-DOPA), a precursor of melanin, contains a cis-diol structure in its catechol skeleton, and excessive melanin production causes skin [...] Read more.

The organogermanium compound 3-(trihydroxygermyl)propanoic acid (THGP) has various biological activities. We previously reported that THGP forms a complex with cis-diol structures. L-3,4-Dihydroxyphenylalanine (L-DOPA), a precursor of melanin, contains a cis-diol structure in its catechol skeleton, and excessive melanin production causes skin darkening and staining. Thus, the cosmetic field is investigating substances that suppress melanin production. In this study, we investigated whether THGP inhibits melanin synthesis via the formation of a complex with L-DOPA using mushroom tyrosinase and B16 4A5 melanoma cells. The ability of THGP to interact with L-DOPA was analyzed by ¹H-NMR, and the influence of THGP and/or kojic acid on melanin synthesis was investigated. We also examined the effect of THGP on cytotoxicity, tyrosinase activity, and gene expression and found that THGP interacted with L-DOPA, a precursor of melanin with a cis-diol structure. The results also showed that THGP inhibited melanin synthesis, exerted a synergistic effect with kojic acid, and did not affect tyrosinase activity or gene expression. These results suggest that THGP is a useful substrate that functions as an inhibitor of melanogenesis and that its effect is enhanced by combination with kojic acid. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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15 pages, 2497 KiB

Open AccessArticle

Effects of Aging on Hair Color, Melanosome Morphology, and Melanin Composition in Japanese Females

by Takashi Itou, Shosuke Ito and Kazumasa Wakamatsu

Int. J. Mol. Sci. 2019, 20(15), 3739; https://doi.org/10.3390/ijms20153739 - 31 Jul 2019

Cited by 19 | Viewed by 7637

Abstract

In a previous study, we showed that the size of melanosomes isolated from Japanese female hairs enlarges with age, and this affects the hair color. In this study, we analyzed the age-dependent changes in hair melanin in order to further explore the factors [...] Read more.

In a previous study, we showed that the size of melanosomes isolated from Japanese female hairs enlarges with age, and this affects the hair color. In this study, we analyzed the age-dependent changes in hair melanin in order to further explore the factors related to hair color changing by aging. A significant positive correlation with age was found in the total melanin amount (TM) and the mol% of 5,6-dihydroxyindole (DHI) units, while no correlation was found in pheomelanin mol%. TM showed significant correlations with hair color parameters and the melanosome volume, suggesting that hair color darkening by aging is caused by the increase in TM due to the enlargement of the size of melanosome. From the measurement of absorbance spectra on synthetic eumelanins with different ratios of DHI and 5,6-dihydroxyindole-2-carboxylic acid (DHICA), we found that the increase in DHI mol% also contributes to the darkening of hair color by aging. In addition, the level of pyrrole-2,3-dicarboxylic acid (PDCA), a marker of DHI melanin, showed a significant negative correlation with the aspect ratio of melanosome, suggesting a contribution of DHI melanin to the change in melanosome morphology by aging. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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18 pages, 3335 KiB

Open AccessArticle

The Oxidative Pathway to Dopamine–Protein Conjugates and Their Pro-Oxidant Activities: Implications for the Neurodegeneration of Parkinson’s Disease

by Kazumasa Wakamatsu, Kenta Nakao, Hitomi Tanaka, Yuki Kitahori, Yui Tanaka, Makoto Ojika and Shosuke Ito

Int. J. Mol. Sci. 2019, 20(10), 2575; https://doi.org/10.3390/ijms20102575 - 25 May 2019

Cited by 16 | Viewed by 4582

Abstract

Neuromelanin (NM) is a dark brown pigment found in dopaminergic neurons of the substantia nigra (SN) and in norepinephrinergic neurons of the locus coeruleus (LC). Although NM is thought to be involved in the etiology of Parkinson’s disease (PD) because its content decreases [...] Read more.

Neuromelanin (NM) is a dark brown pigment found in dopaminergic neurons of the substantia nigra (SN) and in norepinephrinergic neurons of the locus coeruleus (LC). Although NM is thought to be involved in the etiology of Parkinson’s disease (PD) because its content decreases in neurodegenerative diseases such as PD, details are still unknown. In this study, we characterized the biosynthetic pathway of the oxidation of dopamine (DA) by tyrosinase in the presence of thiol peptides and proteins using spectroscopic and high-performance liquid chromatography (HPLC) methods and we assessed the binding of DA via cysteine residues in proteins by oxidation catalyzed by redox-active metal ions. To examine whether the protein-bound DA conjugates exhibit pro-oxidant activities, we measured the depletion of glutathione (GSH) with the concomitant production of hydrogen peroxide. The results suggest that the fate of protein-bound DA conjugates depends on the structural features of the proteins and that DA-protein conjugates produced in the brain possess pro-oxidant activities, which may cause neurodegeneration due to the generation of reactive oxygen species (ROS) and the depletion of antioxidants. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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10 pages, 2260 KiB

Open AccessArticle

Rice Bran Ash Mineral Extract Increases Pigmentation through the p-ERK Pathway in Zebrafish (Danio rerio)

by Yu-Mi Kim, Eun-Cheol Lee, Han-Moi Lim and Young-Kwon Seo

Int. J. Mol. Sci. 2019, 20(9), 2172; https://doi.org/10.3390/ijms20092172 - 02 May 2019

Cited by 11 | Viewed by 3561

Abstract

The purpose of the present study is to evaluate the effect of rice bran ash mineral extract (RBM) on pigmentation in zebrafish (Danio rerio). Melanin has the ability to block ultraviolet (UV) radiation and scavenge free oxygen radicals, thus protecting the [...] Read more.

The purpose of the present study is to evaluate the effect of rice bran ash mineral extract (RBM) on pigmentation in zebrafish (Danio rerio). Melanin has the ability to block ultraviolet (UV) radiation and scavenge free oxygen radicals, thus protecting the skin from their harmful effects. Agents that increase melanin synthesis in melanocytes may reduce the risk of photodamage and skin cancer. The present study investigates the effect of RBM on pigmentation in zebrafish and the underlying mechanism. RBM was found to significantly increase the expression of microphthalmia-associated transcription factor (MITF), a key transcription factor involved in melanin production. RBM also suppressed the phosphorylation of extracellular signal-regulated kinase (ERK), which negatively regulates zebrafish pigmentation. Together, these results suggest that RBM promotes melanin biosynthesis in zebrafish. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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Review

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36 pages, 9632 KiB

Open AccessReview

Chemical Reactivities of ortho-Quinones Produced in Living Organisms: Fate of Quinonoid Products Formed by Tyrosinase and Phenoloxidase Action on Phenols and Catechols

by Shosuke Ito, Manickam Sugumaran and Kazumasa Wakamatsu

Int. J. Mol. Sci. 2020, 21(17), 6080; https://doi.org/10.3390/ijms21176080 - 24 Aug 2020

Cited by 71 | Viewed by 7365

Abstract

Tyrosinase catalyzes the oxidation of phenols and catechols (o-diphenols) to o-quinones. The reactivities of o-quinones thus generated are responsible for oxidative browning of plant products, sclerotization of insect cuticle, defense reaction in arthropods, tunichrome biochemistry in tunicates, production of [...] Read more.

Tyrosinase catalyzes the oxidation of phenols and catechols (o-diphenols) to o-quinones. The reactivities of o-quinones thus generated are responsible for oxidative browning of plant products, sclerotization of insect cuticle, defense reaction in arthropods, tunichrome biochemistry in tunicates, production of mussel glue, and most importantly melanin biosynthesis in all organisms. These reactions also form a set of major reactions that are of nonenzymatic origin in nature. In this review, we summarized the chemical fates of o-quinones. Many of the reactions of o-quinones proceed extremely fast with a half-life of less than a second. As a result, the corresponding quinone production can only be detected through rapid scanning spectrophotometry. Michael-1,6-addition with thiols, intramolecular cyclization reaction with side chain amino groups, and the redox regeneration to original catechol represent some of the fast reactions exhibited by o-quinones, while, nucleophilic addition of carboxyl group, alcoholic group, and water are mostly slow reactions. A variety of catecholamines also exhibit side chain desaturation through tautomeric quinone methide formation. Therefore, quinone methide tautomers also play a pivotal role in the fate of numerous o-quinones. Armed with such wide and dangerous reactivity, o-quinones are capable of modifying the structure of important cellular components especially proteins and DNA and causing severe cytotoxicity and carcinogenic effects. The reactivities of different o-quinones involved in these processes along with special emphasis on mechanism of melanogenesis are discussed. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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37 pages, 4406 KiB

Open AccessReview

From Extraction to Advanced Analytical Methods: The Challenges of Melanin Analysis

by Ioana-Ecaterina Pralea, Radu-Cristian Moldovan, Alina-Maria Petrache, Maria Ilieș, Simona-Codruța Hegheș, Irina Ielciu, Raul Nicoară, Mirela Moldovan, Mihaela Ene, Mihai Radu, Alina Uifălean and Cristina-Adela Iuga

Int. J. Mol. Sci. 2019, 20(16), 3943; https://doi.org/10.3390/ijms20163943 - 13 Aug 2019

Cited by 135 | Viewed by 9925

Abstract

The generic term “melanin“ describes a black pigment of biological origin, although some melanins can be brown or even yellow. The pigment is characterized as a heterogenic polymer of phenolic or indolic nature, and the classification of eu-, pheo- and allo- melanin is [...] Read more.

The generic term “melanin“ describes a black pigment of biological origin, although some melanins can be brown or even yellow. The pigment is characterized as a heterogenic polymer of phenolic or indolic nature, and the classification of eu-, pheo- and allo- melanin is broadly accepted. This classification is based on the chemical composition of the monomer subunit structure of the pigment. Due to the high heterogeneity of melanins, their analytical characterization can be a challenging task. In the present work, we synthesized the current information about the analytical methods which can be applied in melanin analysis workflow, from extraction and purification to high-throughput methods, such as matrix-assisted laser desorption/ionization mass-spectrometry or pyrolysis gas chromatography. Our thorough comparative evaluation of analytical data published so far on melanin analysis has proven to be a difficult task in terms of finding equivalent results, even when the same matrix was used. Moreover, we emphasize the importance of prior knowledge of melanin types and properties in order to select a valid experimental design using analytical methods that are able to deliver reliable results and draw consistent conclusions. Full article

(This article belongs to the Special Issue Melanins and Melanogenesis 2.0: From Nature to Applications)

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